Expression of human immunodeficiency virus type 1 tat from a replication-deficient herpes simplex type 1 vector induces antigen-specific T cell responses

Herpes simplex type-1 virus (HSV-1) based vectors have been widely used in different gene therapy approaches and also as experimental vaccines against HSV-1 infection. Recent advances in the HSV-1 technology do support the use of replication defective HSV-1 as vaccine vectors for delivery of foreign antigens. We have examined the ability of a recombinant replication-defective HSV-1 vector expressing the HIV-1 Tat protein to induce long-term Tat-specific immune responses in the Balb/c murine model. The results showed that vector administration by the subcutaneous route elicits anti-Tat specific T-cell mediated immune responses in mice characterized by the presence of the Tat-specific cytotoxic activity and production of high levels of IFN-gamma.     

Methylene blue improves brain oxidative metabolism and memory retention in rats

Methylene blue (MB) increases mitochondrial oxygen consumption and restores memory retention in rats metabolically impaired by inhibition of cytochrome c oxidase. This study tested two related hypotheses using biochemical and behavioral techniques: (1) that low-level MB would enhance brain cytochrome c oxidation, as tested in vitro in brain homogenates and after in vivo administration to rats and (2) that corresponding low-dose MB would enhance spatial memory retention in normal rats, as tested 24 h after rats were trained in a baited holeboard maze for 5 days with daily MB posttraining injections. The biochemical in vitro studies showed an increased rate of brain cytochrome c oxidation with the low but not the high MB concentrations tested. The in vivo administration studies showed that the corresponding MB low dose (1 mg/kg) increased brain cytochrome c oxidation 24 h after intraperitoneal injection, but not after 1 or 2 h postinjection. In the behavioral studies, spatial memory retention in probe trials (percentage of visits to training-baited holes compared to total visits) was significantly better for MB-treated than saline control groups (66% vs. 31%). Together the findings suggest that low-dose MB enhances spatial memory retention in normal rats by increasing brain cytochrome c oxidase activity.     

New derivatives of pyrrolo[3,4-d]pyridazinone and their anticancer effects

Eighteen new derivatives of pyrrolo[3,4-d]pyridazinone modified at the pyrrole and pyridazine rings were synthesized and 12 of them were evaluated in vitro through anticancer screenings. The structures of new compounds were confirmed by elemental analysis and spectral data (IR, 1H NMR). None of the eight compounds assayed blocked the cell cycle regulating CDK1/cyclin B kinase, whereas two of the six compounds tested were active in anticancer screening at the cell experiments at a concentration of > or = 10(-5) M/l.     

Protein kinase D2 mediates activation of nuclear factor kappaB by Bcr-Abl in Bcr-Abl+ human myeloid leukemia cells

The Bcr-Abl tyrosine kinase activates various signaling pathways including nuclear factor kappaB that mediate proliferation, transformation, and apoptosis resistance in Bcr-Abl(+) myeloid leukemia cells. Here we report that protein kinase (PK) D2, a serine threonine kinase of the PKD family, is a novel substrate of Bcr-Abl. PKD2 was found to be the major isoform of the PKD family expressed in chronic myeloid leukemia cells and is tyrosine phosphorylated by Bcr-Abl in its pleckstrin homology domain. A mutant that mimicks tyrosine phosphorylation of PKD2 in the pleckstrin homology domain activates nuclear factor kappaB independently of its catalytic activity. Furthermore, our data show that Bcr-Abl-induced activation of the nuclear factor kappaB cascade in LAMA84 cells is largely mediated by tyrosine-phosphorylated PKD2. These data present a novel mechanism of Bcr-Abl-induced nuclear factor kappaB activation in myeloid leukemia. Targeting PKD2 tyrosine phosphorylation, not its kinase activity, could be a novel therapeutic approach for the treatment of Bcr-Abl(+) myeloid leukemia.     

Tumor suppressor functions for the Cdk inhibitor p21 in the mouse colon

The Cdk inhibitor p21 regulates p53-mediated growth arrest following DNA damage. It is expressed during epithelial differentiation in a variety of organs including colon. We investigated susceptibility of p21-deficient mice to the colon carcinogen azoxymethane (AOM). After AOM injections, rodents develop putative premalignant lesions called aberrant crypt foci (ACF) that are localized to the distal three centimeters of the colon. p21-deficient mice developed significantly higher numbers of ACF than wild-type mice in response to AOM, and these were not restricted to distal colon. After AOM treatment, increased numbers of lymphoid aggregates were detected in p21-deficient colon. Proliferation was similar in wild type and p21-deficient colon before and after AOM injection, but AOM-induced apoptosis was detected only in wild-type crypt epithelial cells, and not in the p21-deficient colon. The proapoptotic function uncovered for p21 was unexpected, because p21 acts as an inhibitor of apoptosis in many systems, and is not required for p53-dependent apoptosis. Enhanced formation of ACF in p21-deficient mice supports a tumor suppressor function for p21 in the colon. Reduced apoptosis of colon epithelial cells with deleterious mutations may be an initiating event in the formation of ACF, with inflammatory cell cytokine expression contributing to their further expansion.     

Inactivation of HTB63 human melanoma cells by irradiation with protons and gamma rays

The effects of single irradiation with gamma rays and protons on HTB63 human melanoma cell growth were compared. The exponentially growing cells were irradiated with gamma rays or protons using doses ranging from 2-20 Gy. At 48 h of post-irradiation incubation under standard conditions, cell survival and induction of apoptotic cell death were examined. The best effect of the single irradiation with gamma rays was the reduction of cell growth by up to 26% (p=0.048, irradiation vs. control), obtained using the dose of 16 Gy. The same doses of proton irradiation, having energy at the target of 22.6 MeV, significantly inhibited melanoma cell growth. Doses of 12 and 16 Gy of protons provoked growth inhibition of 48.9% (p=0.003, irradiation vs. control) and 51.2% (p=0.012, irradiation vs. control) respectively. Irradiation with 12 and 16 Gy protons, compared to the effects of the same doses of gamma rays, significantly reduced melanoma cell growth (p=0.015 and p=0.028, protons vs. gamma rays, respectively). Estimated RBEs for growth inhibition of HTB63 cells ranged from 1.02 to 1.45. The electrophoretical analyses of DNA samples and flow cytometric evaluation have shown a low percentage of apoptotic cells after both types of irradiation. The better inhibitory effect achieved by protons in contrast to gamma rays, can be explained considering specific physical properties of protons, especially taking into account the highly localized energy deposition (high LET).     

Lipids, lipid peroxidation and 7-ketocholesterol in workers exposed to lead

The study population included healthy men and hypertensive employees of zinc and lead steelworks in the south of Poland. Workers exposed to lead (n=137) were divided into two groups: the first included employees with low exposure to lead (LL) with mean blood lead (PbB) 25-40 microg/dL and the second one with PbB over 40 microg/dL (HL group). The administration workers (n=35) were the control group. Evaluation of lipids and oxidative changes of cholesterol and lipids were estimated in blood samples. No significant changes in concentration of 7-ketocholesterol and blood lipids (cholesterol, HDL, LDL, triglycerides) were found. Lipid peroxidation (LP) was significantly higher in both exposed groups in plasma and in the HL group in erythrocytes when compared with control. There can be two independent sources of LP increase: the first is connected with the direct effect of lead's ions on erythrocytes, the second is the prooxidative effect of delta-aminolevulinic acid. Hypertension in the HL group when compared with people with PbB below 40 microg/dL (OR 4.4, 95%CI 1.4-14.5) was found more often. LP significantly increased by about 71% and concentration of 7-ketocholesterol by about 122% in hypertensives when compared with normotensives in the HL group.     

Changes of axial dimensions of the eye during growth in emmetropia, myopia and hyperopia

PURPOSE: The aim ot this study was to evaluate changes ot axial dimensions ot the eye during growth in emmetropia, myopia and hyperopia. MATERIAL AND METHODS: We examined 183 children (363 eyes) aged 4 to 19 with emmetropia, myopia and hyperopia. All measurements were performed after cycloplegia with 1% tropicamidum (Polfa Warszawa). Total and corneal refraction was examined with autokeratorefractometer (Nikon NRK-8000). Then we used ultrasound biometer Ocuscan (Alcon, USA), to measure axial length of the eye, axial length of the vitreous cavity, axial dimension of the lens and axial depth of the anterior chamber. RESULTS AND CONCLUSIONS: 1. Growth of the axial length of the emmetropic eyes is finished at the age of 12, in hyperopic eyes in the age of 11 and in myopic eyes growth is proportional until the age of 14 and then significantly accelerates. 2. Growth of the axial length is mainly caused by increasing axial length of vitreous cavity. A little role in human eye growth is also played by increasing depth of the anterior chamber. 3. Between 4 and 19 years old, mean cycloplegic axial dimension of the lens is slightly decreasing in emmetropic and hyperopic eyes, whereas in myopic eyes is constant.     

Cyclosporine A and its non-immunosuppressive derivative NIM811 induce apoptosis of malignant melanoma cells in in vitro and in vivo studies

Advanced melanoma is a highly malignant tumor with an increasing incidence that has a poor prognosis due to resistance to common therapeutic strategies. We have demonstrated previously that cyclosporine A (CsA) induces apoptosis of rat glioma cells, reactive astrocytes, and fibroblasts. In our present study, we investigated effects of CsA and its nonimmunosuppressive derivative NIM811 on survival of human and murine melanoma cells. We demonstrated that CsA and NIM811 affect survival of human and murine melanoma cells and induce morphological changes, alterations in nuclear morphology and an internucleosomal DNA fragmentation, consistent with an apoptotic type of death. Western blot analysis showed an activation of caspases 9, 7, 3 and PARP cleavage detectable at 24 hr after exposure of human melanoma cells to the drugs. CsA and NIM811 induced a significant increase in subG1 population of murine B16F10 melanoma cells indicative of apoptotic DNA fragmentation. Studies in murine model of melanoma showed that NIM811, but not CsA, retards tumor progression and significantly decreases tumor volume after intratumoral application. Our findings indicate that CsA and its derivatives may be new candidates for the treatment of melanoma patients.